Food Science Packaging – Michael A. Moskowitz, James K. Liao, Eyal S. Ron, Mary Nallin Omstead, Palmetto Pharmaceuticals LLC
Abstract for “Increasing cerebral bioavailability for drugs”
“A method and compositions for increasing cerebral bioavailability blood-born compositions is provided by administering the composition while increasing brain NO levels. This can be achieved by increasing NO levels by stimulating the production of NO, particularly by administering Larginine. Other agents that increase NO levels independently of ecNOS may also be used. Increased cerebral blood flow means that drugs are also carried into the brain. Increased flow will expose the site of action to more drug molecules. Increased NO production may facilitate administration of drugs not easily absorbed to the brain and/or reduce serum concentrations necessary for desired physiological effects.
Background for “Increasing cerebral bioavailability for drugs”
The blood-brain barrier prevents compounds from freely circulating in the bloodstream of vertebrates from reaching brain tissue. This barrier protects the brain against external influences. This beneficial role can be detrimental if the drug cannot cross the blood-brain border. It may be impossible to give a drug in sufficient doses to raise the blood levels enough to have a beneficial effect on the brain. This is especially true if the drug being administered to the brain causes side effects that are harmful or undesirable to the rest of the body. This problem can be exacerbated when the condition that requires therapy is associated with reduced blood flow to the brain (e.g., ischemic stroke) or a cardiovascular event that results in decreased blood flow or blood pressure to the brain.
Stroke is the third leading cause of death in developed countries. It can be defined as an abrupt impairment of brain function due to a variety pathologic changes. Ischemic strokes account for approximately 80% of strokes. This is due to restricted blood flow. Patients suffering from strokes may benefit from an increase in blood flow and increased delivery of anti-stroke or neuroprotective drugs.
The pathophysiology and causes of cerebral ischaemia have been linked to both apoptotic and excitotoxic mechanisms. MK-801 is a non-competitive NMDA-channel blocker and glutamate antagonist that protects the brain of rats from ischaemic damage. It is also the prototype for neuroprotective drugs that increase resistance to ischemic injuries. Clinical trials of neuroprotectants failed because inadequate blood (brain level) was not possible or because they were toxic.
“Reduced cerebral blood flow could be a factor in the uptake and penetration of lipophilic drugs such as MK-801 into brain tissue. The uptake of these drugs is severely affected by ischemia and reduced blood flow. Stroke can reduce the possibility of reaching therapeutically appropriate brain levels of neuroprotective drug. It is therefore highly desirable to be able to improve cerebral blood flow and drug delivery, particularly under stroke conditions.
“It is the object of this invention that a method be developed to increase cerebral bioavailability for drugs.”
“It is the object of this invention that a method be developed to increase cerebral bioavailability of drugs as a result of increased cerebral blood flow.”
“Nitric dioxide has been proven to act as a vasodilator in the peripheral vasculature of normal tissue. Surprisingly the inventors discovered that increasing NO levels by endothelial and/or non ecNOS dependent mechanisms, or via the generation of nitric dioxide (eNOS), can affect the vasculature of brain tissue. This causes vasodilation, without any loss in blood pressure. The brain vessels release nitric dioxide, which causes blood flow to increase through the brain tissue. This is independent of blood pressure increases. The present invention uses the blood-pressure-independent increase in blood flow through brain tissue to increase cerebral bioavailability of blood-born compositions.”
The present invention allows for greater cerebral bioavailability and brain NO levels of blood-born compositions. This can be achieved by increasing NO levels by stimulating the production of NO, particularly by administering Larginine. Other agents that increase NO levels independently of ecNOS may also be used. Increased cerebral blood flow means that drugs are also carried into the brain. Increased flow will expose the site of action to more drug molecules. Increased NO production may facilitate administration of drugs not easily absorbed to the brain and/or reduce serum concentrations necessary for desired physiological effects.
“The present invention, in an important embodiment, allows for increased delivery of drugs to brain tissues by administering the drug (also known as the?second agent?) or a?physiologically active compound (or agent). While increasing brain NO levels. This can be achieved by increasing NO levels by stimulating the production of NO, particularly L-arginine. It may also be possible to administer agents that increase NO levels without ecNOS or any combination thereof. These agents should be administered in quantities that increase cerebral blood flow and NO levels. Increased cerebral blood flow means that drugs are also carried into the brain with increased blood flow. Increased flow will expose the site of action to more drug molecules. Increased NO production may facilitate administration of drugs not easily absorbed to the brain and/or reduce serum concentrations necessary for desired physiological effects.
“In one preferred embodiment of this invention, the method for enhancing delivery of a desired composition in brain tissue of an individual comprises introducing the composition substantially concurrently with a blood flow enhancing level of L-arginine.”
“In another preferred embodiment of this invention, this invention provides a means to enhance delivery of a desired component to brain tissue of an person. This involves introducing the composition substantially contemporaneously to a blood flow-enhancing amount L-arginine or a bloodflow-enhancing amount non-ecNOS NO generating system.
Agents such as HMG -CoA reductase and rho -GTPase inhibits and inhibitors of actin cytoskeletal organisation are preferred not to be administered in the methods described in the present invention. They are therefore not included in the compositions of the invention. Preferably, protein kinase C and isoquinolinesulfonyl compound or their derivatives, such as H-7 and H-8, are not administered according to the invention. Cyclosporin-A (Csa-A) may not be administered according to certain embodiments.
“The present invention can be used whenever it is desired to increase cerebral bioavailability for a drug. The term subject is defined as humans, animals other than human primates, and dogs, cats. This invention is therefore useful for therapeutic purposes as well as for research purposes, such as testing in animal models or in vitro models for medical, physiological or metabolic conditions.
“Nitric oxide (NO),” has been identified as an uncommon messenger molecule that plays a variety of physiological roles in the cardiovascular, neurologic, and immune systems. (Griffith T M et. al., J Am Coll Cardiol 1988, 12:797-806). It is responsible for blood vessel relaxation, neurotransmission, and pathogen suppression. NO Synthase produces NO from L-arginine’s guanidino nitrogen. (Moncada, S. and Higgs E., Eur J Clin Invest 1991, 21(4), 361-374). At least three types of NO Synthase are known to exist in mammals. The two calcium-dependent isoenzymes of NO Synthase are expressed in neurons (nNOS), endothelial cells, and macrophages (Type III-ecNOS). Endothelial cell Nitric oxide Synthase is the Type III version of the enzyme that can be found in the endothelium. Its reactivity, different pathways of formation and metabolism can explain the many physiological and pathological consequences of NO.
The cerebral blood flow can be increased to increase the drug’s bioavailability in cerebral areas. The cerebral bioavailability can be increased by increasing cerebral blood flow, according to the present inventors.
The present inventors have shown that brain bioavailability, especially lipophilic drugs can be improved by concurrent administration of L-Arginine and other agents that increase NO production by ecNOS or non-ecNOS. These agents should be administered in quantities that increase cerebral blood flow and NO levels. The compositions of the present invention can also be used prophylactically to reduce the risk of stroke, brain injury, or illness. The present invention also allows for the prophylactic, chronic, and acute administration of compositions. This will increase cerebral uptake of drugs to treat stroke or other brain injuries.
L-arginine is a substrate for endothelial Nitric Ox Synthesise (eNOS). L-arginine administration will increase the mass production of nitric oxygen (NO). L-arginine administration causes cerebral blood flow to increase within minutes. A typical increase in cerebral blood flow can be seen within ten to fifteen minute. The maximum increase may be noticed within twenty to sixty minute. Both the infusion rate and the clearance rate determine the time at which maximum effect occurs. Maximum L-arginine concentration at the end will be achieved if the infusion rate and clearance rate are equal.
The present invention allows for greater cerebral bioavailability by administering the drug at interest and increasing cerebral blood flow through increasing brain NO levels. This may be achieved by increasing NO levels by stimulating the production of NO, particularly L-arginine. Other agents that increase NO levels independently of ecNOS can also be used. These agents should be administered in doses that increase cerebral blood flow and cerebral NO levels. Increased cerebral blood flow means that drugs are also carried into the brain via the blood stream. The site of action will become more vulnerable to drug molecules due to increased flow. The stimulation of increased NO production may facilitate administration of drugs to brains, particularly those that are difficult to introduce to brains. In addition, the serum concentration required to achieve desired physiological effects may be decreased.
There are many ways to measure the NO level in cells or tissues. Endothelial dependent relaxation is one phenotypic measure used in the art. This response is affected by NO levels. A nitric oxygen meter can measure the amount of nitric dioxide in a sample. The techniques described above, along with additional techniques, are all well-known to anyone of average skill in the art.
“The invention allows not only to re-establish normal base-line NO levels, but also allows for an increase in NO levels beyond normal base line levels. Normal base-line levels refer to those found in a normal control group. They are adjusted for age and have no symptoms that would indicate an alteration in nitric oxide levels, such as hypoxic conditions or hyperlipidemia. The assay activity used and the age of the participants will determine the actual level. An abnormal situation, such as stroke, can cause nitric oxide levels to drop below normal levels. In cases of stroke, nitric dioxide levels are depressed below their normal levels. Surprisingly the methods and compositions of the invention can restore normal base-line levels in these abnormal conditions. However, nitric dioxide levels can be raised desirably much higher than normal base-line levels. In the context of this invention, “increasing NO levels” means: This includes both normal NO levels being restored and increasing NO levels beyond normal baseline levels.
The invention includes the treatment of ischemic stroke. Ischemic stroke (ischemic brain infarction) refers to an acute neurologic injury caused by a decrease of blood flow to the brain’s blood vessels. There are two main types of ischemic stroke: thrombotic or embolic.
“An unexpected finding was made regarding the treatment of an ischemic stroke. The invention’s treatment can improve blood flow to the brain even after an ischemic stroke. The present invention led to cerebral blood flow being greater in the treated animals than the control. The increase in nitric dioxide levels is thought to be responsible for the positive results.
“The invention’s most important embodiment is the treatment of subjects at high risk for ischemic stroke. Subjects with an abnormally high risk of having an ischemic stroke as defined herein are those who have been identified by conventional medical practice as having known stroke risk factors or increased risk of developing cerebrovascular events. The risk factors for cardiac disease are similar to those associated with stroke. The primary risk factors include hypertension, hypercholesterolemia, and smoking. Other risk factors include atrial fibrillation and recent myocardial injury.
Subjects at abnormally high risk of an ischemic attack include those who are undergoing diagnostic or surgical procedures that could result in emboli being released, lowering blood pressure, or decreasing blood flow to their brains. These procedures may include brain angiography, carotid endarterectomy and brain angiography. Individuals with abnormally high risk of an ischemic attack include those who have a cardiac condition that could cause decreased blood flow to their brains, such as atrial fibrillation or ventrical tachycardia, dilapidation, and any other conditions that require anticoagulation. Individuals with abnormally high risk of an ischemic attack include those who have conditions such as arteriopathy, brain vasculitis, and congenital blood vessel diseases such as CADASIL syndrome. Migraines are also considered to be at increased risk. “In certain embodiments, the subject does not have a high level of hypercholesterolemic, hypertriglyceridemic, or both.
The treatment of stroke according this invention is for stroke victims who have had a stroke. It can also be used as a prophylactic treatment. Prophylactic treatment for short term is recommended for patients who have undergone diagnostic or surgical procedures that could result in emboli release, lowering blood pressure, or decreases in blood flow to their brains. This will reduce injury from any ischemic events. Patients with cardiac conditions, which may result in decreased blood flow to their brains, or conditions directly affecting the brain’s vasculature, should consider long-term or chronic prophylactic therapy. If the treatment is prophylactic, it will be for those who are at an abnormally high risk of having an ischemic stroke. Acute treatment is possible for subjects who have suffered a stroke. The administration of acute treatment to stroke subjects is preferably at the onset or a significant change in symptoms.
“Another important aspect of the invention is the treatment of subjects suffering from a neurodegenerative disorder. “Neurodegenerative disease” is a broad term. Any pathological condition involving neuronal degradation, such as Huntington’s Disease or Parkinson’s Disease (or ALS), is included in the term “neurodegenerative disease”. Preferably, the neurodegenerative condition is Alzheimer’s Disease. Alzheimer’s Disease, a progressive neurodegenerative disorder, is characterised by the death and loss of function of nerve cells in various areas of the brain. This causes cognitive decline such as memory loss and language loss. Although the cause of nerve cell death remains unknown, the cells can be identified by unusual helical protein filaments (neurofibrillary knots) in nerve cells. Also, degeneration is seen in cortical brain regions, particularly the frontal and temporal. The present invention can increase cerebral bioavailability for suitable drugs. This is especially beneficial to patients suffering from neurodegenerative diseases such as Alzheimer’s.
The present inventors have shown that brain bioavailability, especially lipophilic drugs can be improved by concurrent administration of L-Arginine and other agents that increase NO production by ecNOS or non-ecNOS. These agents should be administered in sufficient quantities to increase cerebral blood flow and brain NO levels. The compositions of the invention can also be used prophylactically to reduce the risk of stroke, other brain injuries, or illnesses. The present invention also promotes cerebral uptake of drugs to treat stroke and other brain injuries.
The present invention allows for greater cerebral bioavailability by administering the drug and increasing cerebral blood flow through the increase in NO levels. This may be achieved by increasing NO levels by increasing eNOS production, particularly by administering Larginine. Other agents that increase NO levels independently of ecNOS can also be used. These agents should be administered in sufficient quantities to increase blood flow to the brain and/or NO levels. Increased cerebral blood flow means that drugs are also carried into the brain with increased blood flow. The site of action will become more vulnerable to drug molecules due to increased flow. The stimulation of increased NO production may facilitate administration of drugs to brains, particularly those that are difficult to introduce to brains. In addition, the serum concentration required to achieve desired physiological effects may be decreased.
An agent (preferably Larginine) that increases the ecNOS levels by preexisting and/or non-ecNOS NO generation is administered to the subject who requires enhanced drug delivery. An agent that increases ecNOS by preexisting (preferably L-arginine), and/or a system that does not generate NO may be called a?NO-increasing combination. Each of these agents can be called an ‘NO-increasing agency. These agents could be combined to be called?NO-increasing agent.
“As used in the present application, the terms “NO-increasing cocktail?” a?NO-increasing agent? Agents such as HMG?CoA reductase inhibits, rho?GTPase inducers and inhibitors for actin cytoskeletal order are not included. These agents are described in detail in WO 99/18952, WOM 99/47153 and WOM 00/03746, which have been incorporated herein by reference. The terms “NO-increasing cocktail” and “NO-increasing agent(s)” are used throughout this application. a?NO-increasing agent? Do not contain protein kinase C inhibitors, Cyclosporin-A (Csa-A), or Isoquinolinesulfonyl compound or their derivatives, such as H-7 or H-8.
The NO-increasing agent/mixture is administered substantially simultaneously with the second agent (the drug whose delivery will be improved). The preferred embodiment of the NO-increasing cocktails includes L-arginine, at least one nonecNOS NO generating system, and L-arginine.
“The components of the NO -increasing cocktail should be administered substantially simultaneously with each other. As we will see, “substantially contemporaneous administration” is a term that can be used to describe a combination of components. It should be understood broadly and includes many types of administration. Referring to the components of NO-increasing cocktails, “substantially contemporaneously?” This means that the components are administered in such a way that they work together to produce a significant increase in NO levels in the subject or in particular tissues.
To be considered a “NO-increasing cocktails,” agents not only need to be administered in one form or one unitary dose, but also in multiple doses. “NO-increasing cocktails” does not mean that agents used in the present invention must be administered in a single formulation or in one unitary dosage.
“To stimulate higher NO production by ecNOS an?ecNOS activating ingredient (or agent) is required. is administered to the subject who requires enhanced drug delivery. Agents that increase NO production through ecNOS Agents that increase NO production through preexisting ecNOS. These terms can also be used interchangeably to denote ecNOS activating agent. These agents are also known as agents which increase NO production via preexisting EcNOS. This does not mean that these agents can’t also increase NO production of ecNOS produced during or after administration. Instead, the term “preexisting” is used. This is to show that these agents don’t increase or downregulate the expression ecNOS.
The activating component must be administered concurrently with the drug whose delivery is being enhanced. An eNOS substrate such as L-arginine is a preferred component that activates eNOS. This increases NO production. Alternative eNOS activating components include cofactors of eNOS, such as NADPH or tetrahydrobiopterin.”
“Compounds that increase NO production by preexisting ecNOS can do so through a variety of mechanisms. L-arginine and other ecNOS substrates increase NO production by mass action. NADPH and Tetrahydrobiopterin are cofactors that increase NO production by increasing the ability for ecNOS catalyze the substrate to NO conversion. These ecNOS substrates, e.g. L-arginine and cofactors (e.g. NADPH, Tetrahydrobiopterin etc.) may be used as ecNOS substrates. Natural or synthetic, they can be either natural or artificial. “Compounds that increase NO production by preexisting NOS can act in a cooperative, additive, or synergistic manner with agents that increase NO levels through non-ecNOS dependent mechanisms (i.e. non-ecNOS no-generating systems).
L-arginine is a substrate for endothelial Nitric Ox Synthesise (ecNOS). L-arginine administration will increase the mass production of nitric oxygen (NO). L-arginine administration causes cerebral blood flow to increase within minutes. A typical increase in cerebral blood flow can be seen within ten to fifteen minute. The maximum increase can occur within twenty to sixty minute. Both the infusion rate and the clearance rate determine the time at which maximum effect occurs. Maximum L-arginine concentration at the end will be achieved if the infusion rate and clearance rate are equal.
“EcNOS activating components” is the term used in the present application. HMG-CoA reductase inhibits, rho?GTPase inducers and inhibitors of actin-cytoskeletal organization are not included. These agents are described in detail in WO 99/18952, WOM 99/47153 and WOM 00/03746, which have been incorporated herein by reference. The term “ecNOS activating constituent” is used throughout this application. Does not include protein kinase C inhibits, cyclosporin A (Cs-A), isoquinolinesulfonyl compound or their derivatives. These are not administered according to the invention and are not included within the compositions according the invention.
When administered to biological systems, “NO donors” are compounds that release or produce NO, or have NO-related activity. Feelisch (1998) Naunyn Schmiedebergs Arch Pharmacol 358(1): 113-22. There are many examples of NO donors that are well-known and will be discussed in detail below. Subjects can also increase NO levels by inhaling NO.
“Non-ecNOS NO generating devices directly increase NO levels in a subject, tissue, or cell without relying upon ecNOS and other Nitric Oxide synthases. The invention defines “non-ecNOS” NO-generating systems as: Compounds that, when administered to a subject increase NO levels without relying upon ecNOS. Inhalation of NO can be used to administer NO directly to the subject. A patient may also receive NO via administration of NO donors. “Non-ecNOS NO generating systems” includes both NO and NO donors.
“The term “non-ecNOS no-generating systems” is used throughout this application. HMG-CoA reductase inhibits, rho?GTPase inhibitions and inhibitors for actin cytoskeletal organisation are not included. These agents are described in detail in WO 99/18952, WOM 99/47153 and WOM 00/03746, which have been incorporated herein by reference. The term “non-ecNOS no-generating systems” is used throughout this application. Does not include protein kinase C inhibitors (Cs-A), cyclosporin A (Cs-A), isoquinolinesulfonyl compound or their derivatives. These are not administered according to the invention and are not included within the compositions according the invention.
“It is possible to optimize routinely the administration of inhaled NO for the use in the compositions and methods of the present invention.” The properties and activities of NO inhaled are well-known. Administration of inhaled NO to human subjects is described at least in Hoeper, et al., Abman, et al., Carrier, et al., Kinsella, et al., and Kuhlen, et al. See, e.g., Hoeper, et al. (1999) Respir Med. 93(1): 62-4; Abman, et al. (1994) J. Pediatr. 124(6): 881-8; Carrier, et al. (1999) 18(7): 664-7; Kinsella, et al. (1993) J. Pediatr. 122(5Pt 1): 803-6; Kuhlen, et.al. (1999) Intensive Care Med. 25(7): 752-4. The texts of these publications are incorporated by reference herein.
Multiple groups have described the chemical kinetics and methods of reducing NO2 buildup in systems for inhalation of NO2. See, e.g., Tsukahara, et al. (1999) Nitric Oxide (3): 191-8. Lindberg, et. al. (1998) Br. J Anaesth 80(2) 213-7. The texts of these publications are included herein as a reference. Multiple groups have also provided detailed descriptions of techniques for monitoring and delivery of inhaled NO. See, e.g., Kirmse, et al. (1998) Chest 113(6): 1650-7; Shibata (1996) Acta Paediatr Jpn 38(2): 143-6; Young, et al. 1996 Intensive Care Med. 22(1):77-86; Hess, and al. Respir Care Clin N Am 3, 3(3), 371-410. The texts of these publications are included herein as a reference. The art describes technical considerations such as NO gas concentrations. See, e.g., Kinsella, et al. (1999) Curr Opin Pediatr 11(2): 121-5; Foubert, et al., (1999) Anaesthesia 54(3): 220-5; Breuer, et al. (1997) Eur. J. Pediatr. 156(6): 460-2; Moon, et al. (1997) Biomed Instrum Technol 31.2: 164-8; Hart (1999) Chest 115.5: 1407-17. The texts of these publications are included herein as a reference.
“As used herein, the term ‘NO donors? A large group of molecules that have varying properties but all release or produce NO, or are related to NO, when applied to biological systems. Feelisch (1998) Naunyn Schmiedebergs Arch Pharnacol 358,(I): 113-22. The text of that publication is included herein as a reference. These compounds are well-known in art. Examples of NO donors include nitroglycerin, nitric oxide/nucleophile adducts (NONOates), including diethylamine/NO complex sodium (Dea/NO) and spermine/NO complex sodium; S-nitrosothiols, also called NO+ equivalents, such as S-nitroso-L-glutathione (GSNO) and S-nitroso-N-acetyl-D,L-penicillamine (SNAP); nitrosylated proteins, such as nitrosylated bovine serum albumin (BSA). See, e.g., Ewing, et al., (1997) J. Pharmacol. Exp. Ther. 283(2):947-54; Vidwans, et al. (1999). J. Neurochem 72(5) : 947-54; Vidwans, et al. SPM-5185, an organic cysteine-containing drug, undergoes a biotransformation process that releases NO when it is exposed to physiological conditions. See, e.g., Vinten-Johansen, et al. (1995) Int. J Cardiol. 50(3): 273?81. The text of that publication is incorporated by reference herein. Another NO donor that has been therapeutically used in humans is sodium nitrosprusside. See, e.g., Thomas, et al. (1999) Neurosurgery 44(1):48-57, 57-8; Thomas, et al. (1999) Stroke (30(7): 1409-16. The texts of these publications are included herein by reference. The heterocyclic NO-releasing compound, which includes mesionic heterocycles such as SIN-1 and heterocyclic NOs such as furoxane carbboxamides, are other examples of NO donors. See, e.g., Schonafinger (1999) Farmaco 54(5): 316-20 and Hou, et al., (1999) Curr. Pharm. Des. 5(6): 417-441. The texts of these publications are incorporated by reference. U.S. Patent. Describes additional NO donors. No. 5,910,316 to Keefer, et al., U.S. Pat. No. 5,525,357 to Keefer, et al., U.S. Pat. No. Loscalzo, et al. and U.S. Pat. No. No.
It is routine optimization for a skilled worker in the art to choose NO donors that are suitable for the compositions and methods according to the invention. A skilled worker in the arts can also routinely optimize dosages and administration methods to suit the compositions and techniques of the invention. The activities and properties of NO donors have been well-known. Schmidt et al. have created a mathematical model to predict the NO concentrations of donor compounds over time. Schmidt, et al. Schmidt, et al. Morley and Keefer discuss in depth NONOates. Kal, et. al. describes in detail how nitroglycerin is administered to patients. Morley, et al., (1993) J. Cardiovasc. Pharmacol. 22 Suppl. 7: S3-9; Kal, et al. (1999) Anesth. Analg. Analg.
“Estrogens, ACE inhibitors and other hormones can also increase NO levels. While estrogens and ACE inhibitors can be used in the compositions and methods according to the invention, they are not included under the term?agents that increase NO production by preexisting ecNOS? ?NO-increasing compound,? ?NO-increasing cocktail,? ?non-ecNOS NO-generating system,? or ?ecNOS activating component.? Estrogens can be described as a specific group of molecules that are well-known to those with ordinary skill in the field. We won’t go into detail about them. They share high levels of structural similarity. ACE inhibitors have also been well characterized. However, they don’t always share structural homology.
Angiotensin converting enzyme (or ACE) is an enzyme that catalyzes angiotensin I and II conversions. ACE inhibitors are amino acids, peptides (including di and tri peptides), and antibodies to ACE. They interfere in the renin?angiotensin systems by inhibiting the activity ACE and thereby reducing/eliminating the formation of the pressor substance angiotensin 2. ACE inhibitors are used medically to treat hypertension and congestive heart disease, as well as myocardial injury, myocardial damage, renal disease, and congestive heart failure. ACE inhibitors are acylmercapto, mercaptoalkanoyl proslines like captopril (U.S. Patent. No. No. No. No. No. 4,374,829), lisinopril (U.S. Pat. No. 4,374,829), quinapril (U.S. Pat. No. No. No. No. No. No. No. No. No. 4,410,520), phosphinylalkanoyl proslines such as fosinopril. (U.S. Pat. No. No.
“In some embodiments, the second drug (i.e. the drug that is sought to enhance delivery) is administered to a subject suffering from a condition. The second agent is administered in sufficient amounts to treat the condition. This increases blood flow to the subject’s tissues.
The second agent can be any pharmacological or diagnostic compound. Agents with a brain-specific site of action are preferred second agents. These agents can be analgesics, anesthetics, anti-andrenergic agents, amino acids and antagonists, as well as antidote (anti-anxiety agent), anticolvunsant/antidepressant/antihypertensive/antihypertensive/antifibrinolytics, antihypertensive/antihypertensive/antiparkinsonian agent, antiobessional agent or antiparkins, neuromuscular block, neuroprotective, and neuromuscular blockers, neuromuscular blockers, and neuroprotective, serotonin agonists, and calcium-channels. NMDA antagonist, post-stroke and post-head trauma treatment, psychotropic, sedative, sedative-hypnotic, selective serotonin uptake inhibitor, serotonin inhibitor, tranquilizer, and treatment of cerebral ischemia, calcium channel blockers, free radical scavengers-antioxidants, GABA agonists, glutamate antagonists, AMPA antagonists, kainate antagonists, competitive and non-competitive NMDA antagonists, growth factors, opioid antagonists, phosphatidylcholine precursors, serotonin agonists, sodium- and calcium-channel blockers, and potassium channel openers.”
“In addition to the foregoing brain-specific categories of agents, examples of categories of other pharmaceutical agents that can be used as second agents include: adrenergic agent; adrenocortical steroid; adrenocortical suppressant; alcohol deterrent; aldosterone antagonist; amino acid; ammonia detoxicant; anabolic; analeptic; analgesic; androgen; anesthesia, adjunct to; anesthetic; anorectic; antagonist; anterior pituitary suppressant; anthelmintic; anti-acne agent; anti-adrenergic; anti-allergic; anti-amebic; anti-androgen; anti-anemic; anti-anginal; anti-anxiety; anti-arthritic; anti-asthmatic; anti-atherosclerotic; antibacterial; anticholelithic; anticholelithogenic; anticholinergic; anticoagulant; anticoccidal; anticonvulsant; antidepressant; antidiabetic; antidiarrheal; antidiuretic; antidote; anti-emetic; anti-epileptic; anti-estrogen; antifibrinolytic; antifungal; antiglaucoma agent; antihemophilic; antihemorrhagic; antihistamine; antihyperlipidemia; antihyperlipoproteinemic; antihypertensive; anti-infective; anti-infective, topical; anti-inflammatory; antikeratinizing agent; antimalarial; antimicrobial; antimigraine; antimitotic; antimycotic, antinauseant, antineoplastic, antineutropenic, antiobessional agent; antiparasitic; antiparkinsonian; antiperistaltic, antipneumocystic; antiproliferative; antiprostatic hypertrophy; antiprotozoal; antipruritic; antipsychotic; antirheumatic; antischistosomal; antiseborrheic; antisecretory; antispasmodic; antithrombotic; antitussive; anti-ulcerative; anti-urolithic; antiviral; appetite suppressant; benign prostatic hyperplasia therapy agent; blood glucose regulator; bone resorption inhibitor; bronchodilator; carbonic anhydrase inhibitor; cardiac depressant; cardioprotectant; cardiotonic; cardiovascular agent; choleretic; cholinergic; cholinergic agonist; cholinesterase deactivator; coccidiostat; cognition adjuvant; cognition enhancer; depressant; diagnostic aid; diuretic; dopaminergic agent; ectoparasiticide; emetic; enzyme inhibitor; estrogen; fibrinolytic; fluorescent agent; free oxygen radical scavenger; gastrointestinal motility effector; glucocorticoid; gonad-stimulating principle; hair growth stimulant; hemostatic; histamine H2 receptor antagonists; hormone; hypocholesterolemic; hypoglycemic; hypolipidemic; hypotensive; imaging agent; immunizing agent; immunomodulator; immunoregulator; immunostimulant; immunosuppressant; impotence therapy adjunct; inhibitor; keratolytic; LNRH agonist; liver disorder treatment; luteolysin; memory adjuvant; mental performance enhancer; mood regulator; mucolytic; mucosal protective agent; mydriatic; nasal decongestant; neuromuscular blocking agent; neuroprotective; NMDA antagonist; non-hormonal sterol derivative; oxytocic; plasminogen activator; platelet activating factor antagonist; platelet aggregation inhibitor; post-stroke and post-head trauma treatment; potentiator; progestin; prostaglandin; prostate growth inhibitor; prothyrotropin; psychotropic; pulmonary surface; radioactive agent; regulator; relaxant; repartitioning agent; scabicide; sclerosing agent; sedative; sedative-hypnotic; selective adenosine Al antagonist; serotonin antagonist; serotonin inhibitor; serotonin receptor antagonist; steroid; stimulant; suppressant; symptomatic multiple sclerosis; synergist; thyroid hormone; thyroid inhibitor; thyromimetic; tranquilizer; treatment of amyotrophic lateral sclerosis; treatment of cerebral ischemia; treatment of Paget’s disease; treatment of unstable angina; uricosuric; vasoconstrictor; vasodilator; vulnerary; wound healing agent; xanthine oxidase inhibitor.”
“Throughout this application by terms such as “substantially contemporaneous Administration”,? ?co-administration,? ?substantially contemporaneously,? ?substantially concurrently? It is intended that all of the administered compounds are administered to the subject in a relative order so that they may exert an additive, or even synergistic effect on the subject. On increasing NO levels or delivering a second agent via increased blood flow to a tissue. These terms can be interchanged.
“Substantially contemporaneous Administration? “?Substantially contemporaneous administration” refers to administration of a combination of drugs for increasing NO levels (as discussed herein), such that cerebral blood flow is increased while the second medication is in sufficient serum concentration (i.e. enough to allow the second drug have a physiological effect).
“With respect to NO-increasing agent and components of the NO?increasing cocktail??substantially contemporaneously?” This means that the timing of administration of components is coordinated in such a way that a significant increase in NO levels is achieved in the subject or in particular tissues.
“Substantially simultaneous administration?” This includes administration of agents (both NO -increasing agent and second agents) in one formulation or as a unitary dose. Substantially simultaneous administration This includes agents administered in different dosage formats, formulations, and at different times. As long as the criteria are met for substantially simultaneous administration (as noted above),
“For example, a combination of drugs for increasing NO levels and the second drug could be prepared for intravenous administration. Infusion in one pharmaceutical composition. This allows for simultaneous infusion of a combination that increases NO levels and the second drug. It is preferable that the drug to which the delivery is being increased is in the bloodstream while NO levels are increasing.
The two components can be combined in one oral composition if the second drug is absorbed into bloodstream at a rate similar to that of an oral combination for increasing NO levels (as discussed herein). The pharmacokinetics for the second drug may not be as effective for several hours. If this happens, a combination for increasing NO levels (as discussed herein) can be administered in substantially contemporaneous fashion. This means that the blood flow will increase once the serum concentrations of both drugs have been reached. A combination of drugs for increasing NO levels, as described herein, that are substantially contemporaneous in administration to another drug, whether one or both of them are administered via a nasal, topal, or rectal route or intramuscularly or undercutaneously, is routine for those who are skilled in pharmacology or clinical medicine.
Effective amounts are given to agents (preferably Larginine), which increase NO production by preexisting NO generating systems and/or ecNOS NO generating system. An effective amount is generally any amount that causes an increase in blood flow to or from the brain. This is usually a amount that is effective in increasing NO levels in the brain.
The second or additional agents can also be administered in effective doses. An effective amount of any pharmaceutical preparation is the amount that produces the desired response, whether it’s taken alone or in combination with other agents. It may be a temporary slowing down of the disease’s progression, but it is more effective if the disease is stopped permanently. Routine methods can monitor such results.
The effectiveness of a second agent will depend on how it is administered. The effective amounts of second agent are known or can be easily determined by those skilled in clinical medicine and pharmacology. The present invention aims to decrease the amount of second agents required to achieve a desired effect. Therefore, the effective amounts can be modified by administering the second agent according to the methods of this invention. The new effective amounts can be determined by routine experimentation of those who are skilled in clinical medicine and pharmacology.
The effective doses of both the NO-increasing and second agents will vary depending on the patient’s condition and their individual parameters, including their age, weight and size, as well as the type of concurrent therapy and other factors that the health practitioner has access to. Some forms of administration such as intravenous injections can result in lower doses. If a subject’s response is not sufficient at the initial doses, they may need to be administered higher doses or a more localized route. To achieve the appropriate systemic levels, multiple doses are possible per day.
“It is preferred that the maximum dose of NO-increasing agent or cocktail be used. This is the safest dose according to sound medical judgement. However, those with ordinary skill in the arts will understand that patients may request a lower or more tolerable dose for psychological or medical reasons.
“The agents that increase NO production by preexisting ecNOS, NO, NO donors and other compounds useful according to the invention may be combined, optionally, with a pharmaceutically-acceptable carrier. The term ?pharmaceutically-acceptable carrier? The term “pharmaceutically-acceptable carrier” is used herein to refer to any combination of solid or liquid fillers, diluting agents or encapsulating substances that are suitable for administration into a person. “Carrier” is a term that refers to an organic or inorganic ingredient. The term “carrier” refers to an organic or inorganic component, natural or synthetic, that is used to combine the active ingredient for the purpose of application. All components of the pharmaceutical compositions are also capable of being mixed with the molecules of this invention and with each other in a way that does not substantially impair their pharmaceutical efficacy.
“The pharmaceutical compositions could contain appropriate buffering agents such as acetic acid, citric acid and salt; boric acids in salt; or phosphoric acid.
“The pharmaceutical compositions may also contain, optionally: benzalkonium chloride, chlorobutanol, parabens, and thimerosal.”
There are many routes of administration. There are many options available. The mode of administration will depend on the drug being used, the severity of the condition, and the dose required to achieve therapeutic efficacy. Medically acceptable means that the methods can be used to produce effective levels of active compounds with no clinically unacceptable side effects. These modes of administration include oral, rectal and topical, nasal, interdermal or parenteral. Parenteral is a broad term. It can be subcutaneous, intravenous or intramuscular. For prophylaxis and long-term treatment, intravenous and intramuscular routes may not be suitable.
The pharmaceutical compositions can be conveniently presented in a unit dosage form. They may also be prepared using any of the well-known methods in pharmacy. All methods involve the addition of the active agent(s), or any combination thereof, to a carrier. The compositions are generally prepared by bringing the active compounds into contact with a liquid carrier or a finely divided solid carrier. If necessary, the product can be shaped.
“Compositions that are suitable for oral administration can be presented in discrete units such as tablets, capsules, or lozenges. Each unit contains a predetermined amount. You can also make suspensions from aqueous or non-aqueous liquids, such as syrups, elixirs or emulsions.
“Compositions that are suitable for parenteral administration consist of a sterile, aqueous preparation containing a NO-increasing or cocktail. It should be preferably isotonic with blood of the recipient. This composition may also contain the second or additional agents. They may also be administered separately, but in a substantially parallel fashion. The aqueous preparation can be prepared according to well-known methods, using appropriate dispersing or wetting agent and suspending agents. The sterile injectable preparation also may be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example, as a solution in 1,3-butane diol. Water, Ringer’s solution and isotonic chloride solution are acceptable solvents. Sterile, fixed oils can also be used as a solvent and suspending medium. Any bland, unscented oil can be used for this purpose, including synthetic mono- or di-glycerides. You can also use fatty acids like oleic acid in the preparation injectables. Carrier formulation suitable for oral, subcutaneous, intravenous, intramuscular, etc. Remington’s Pharmaceutical Sciences, Mack Publishing Co. Easton, Pa. has the administrations.
“Other delivery methods include delayed release, time-release and sustained release. These systems allow for the patient to be administered once and then stopped. This increases convenience for both the subject as well as the physician. There are many types of release delivery systems that are known to anyone with ordinary skill in the arts. They include polymer base systems such as poly(lactide-glycolide), copolyoxalates, polycaprolactones, polyacrylates, polyesteramides, polyorthoesters, polyhydroxybutyric acid, and polyanhydrides. U.S. Pat. describes microcapsules containing the above-mentioned polymers that contain drugs. No. 5,075,109. Non-polymer delivery systems include: lipids, including cholesterol esters and neutral fats like mono-di and tri-glycerides, hydrogel release systems, cellusics, sylastic system; peptide-based systems; wax coatings and compressed tablets with conventional binders or excipients. Examples include, but not limited to: (a. erosional systems where the active compound is contained within a matrix like those described in U.S. Pat. Nos. Nos. Nos. Nos. Pump-based hardware delivery systems are also available, some of which can be adapted for implant.
A long-term sustained-release implant might be desirable. The term “long-term release” is used herein to mean that the implant has been constructed and designed to deliver therapeutic levels of active ingredient for at most 30 days and preferably 60. Long-term sustained-release implants are well-known to anyone with ordinary skill in medicine and include some of these release systems.
Pre-packaged combinations of NO increasing agent or agents and NO-increasing cocktail, second agents, or any combination thereof, are preferred embodiments. Combinations of NO-increasing agent/ agents, NO-increasing cocktail, second agents and any combination thereof that are prepackaged using “Blow/Fill/Seal” are even more preferred. technology, as described below.”
“Blow-Fill Seal technology” is a manufacturing process that allows for simultaneous filling and sealing of containers using one machine. Sometimes called form-fill-seal. This process has many advantages over traditional aseptic filling preformed plastic containers (or other types) by eliminating the need for multiple steps. It can also be performed in one machine in a sterile environment and requires very little operator intervention. (J. R. Sharp.?Manufacture Sterile Pharmaceutical Products Using ‘Blow-Fill? Seal? Technology?, Pharm. J., 239, 106 (1987). F. Leo?Blow/Fill/Seal Aseptic Package Technology in Aseptic Pharmaceutical Tech for the 1990’s?. Interpharm Press. Prairie View, Ill. 1989, pp. 195-218).”
The manufacturing process is broken down into several stages. In stage 1, the polyethelene resin undergoes high pressure and temperature, and is then extruded continuously to form a tube. This is known as a parison. The mold is closed when the tube is the right length and the parison cut. The top of the parison can be held in place by securing the bottom. The mold is then moved to the position where it can be filled with sterilized air. The second stage involves the inserting the blow-fillnozzle into the parison, until it seals with the mold’s neck. By blowing compressed air through the parison and expanding it against the walls of a mold cavity, the container is created. The compressed air is then ejected from the container, and a sterile product is poured into the container via the fill nozzle. The nozzle can be retracted back to its original position after the container has been filled. Semi-molten is reached at this stage of the cycle (stage 3). To form the top, separate sealing molds are used to seal the container. The mold is then opened in stage 4. The mold is removed from the machine and the container is filled, sealed, and returned to the origin point to begin the next cycle. You can add additional products to the container anytime during or prior to use through an injection port.
“A preferred embodiment contains a NO-increasing cocktail or agent and a second agent. It is packaged in a Blow/Fill/Seal containers according to the methods herein. Another preferred embodiment is that a NO-increasing cocktail or agent is prepackaged into a Blow/Fill/Seal containers according to the methods herein. A second agent or agents can be injected into this container before or during administration to patients. The second agent may be administered by another method such as injection, oral administration (sublingual administration), inhalation or the like.
“In all of the above embodiments it is preferable that L-arginine be the NO-increasing agents or that L-arginine is the NO-increasing cocktail.”
“In such prepackaging embodiments the NO-increasing agents or cocktails included in these prepackaging embodiments will preferably be sufficient to increase blood flow and NO levels in the brain of the subject. Prepackaged formulations will contain sufficient second agents to achieve the desired effect. It is especially important that the amount of the second agent be adjusted to account for the increased blood flow caused by the NO-increasing agents or cocktails. The effectiveness of each second agent will differ.
“It will be obvious to the skilled artisan, that the considerations in formulating pharmaceutical preparations, and determining the modes of administration for non-ecNOS-generating systems, are similar to those involved in such formulation, determination for agents which upregulate the expression of ecNOS, and compounds which increase NO production by preexisting, ecNOS.”
“Publications WO 99/18952, WO99/47153, WO 00/03746, and WO 00/03746, provide more detailed information about endothelial nitricoxid synthase and their regulation, as well methods of formulating compounds that affect eNOS such as L-arginine and the physiologically active compositions. These publications are herein incorporated as reference.”
“EXAMPLES”
Below are a few examples that will help you better understand the invention. These Examples are only examples and do not limit the scope of the invention.
“Example 1”
“Effects of L-arginine upon Cerebral Blood Flow.”
Infusion of L-arginine at 300 mg/kg i.v. caused moderate (10%) and variable elevations (RCBF), after infusion in several preliminary studies (n=4, data unpublished). The present experiments showed that simvastatin (2 mg/kg) or 450 mg/kg of saline were infused at a steady rate of 100 microliter/kg/min for 15 minutes to wild type mice, mutant mice lacking endothelial oxide synthase, and mice who had been given simvastatin (2 mg/kg) daily. Regional cerebral blood flow (rCBF) was monitored by laser-Doppler flowimetry in groups of urethane-anesthetized, ventilated mice. Other physiological variables, such as mean arterial blood pressure (MABP), heart beat, blood pH, and PaO2 were also observed in the mice.
“Results”
“Physiological variables during laser-Doppler flowimetry in urethane-anesthetized ventilated wild type, simvastatin-treated and eNOS null mice infused with L-arginine or saline are shown in Table 1. In parenthesis, the number of mice in each group are shown. The values are reported as the mean +/-? SEM. SEM. MABP is the mean arterial blood pressure. Sim denotes mice who have been chronically administered simvastatin.
“There were no differences between the groups in heart rate and mean arterial blood pressure during observation, but these values were higher in eNOS-null mice, as previously reported. PaCO2 values did not differ between the two time points of all groups or between-groups, but pH values were lower after L-arginine infusion.
“rCBF Response for L-arginine”
“FIG. “FIG. In parenthesis, the number of mice in each group are indicated. An error bar denotes standard error of mean (SEM) and an asterisk(*) denotes statistically significantly difference (P0.05) when compared to baseline control using one-way ANOVA followed Fisher’s protected least squares difference test.
“L-arginine infusion (450 mg/kg, i.v.) As shown in FIG. 1 (FIG. 1). At 5-10 minutes, the RCBF increased and reached statistical significance 10-15 minutes after infusion. Maximum values reached at 20-25 minutes were 26% higher than the control level, and then they decreased to their normal levels. L-arginine, however, did not increase the rCBF of eNOS null mice. These mutants had values ranging from?4 up to +5% over the 40-minute recording period. In wild-type mice, saline infusion did not significantly increase rCBF.
“rCBF Response To L-arginine Plus Simvastatin.”
“FIG. “FIG. In parenthesis, the number of mice in each treatment group is indicated. sim denotes simvastatin. An error bar denotes SEM, and an asterisk (“*”) denotes statistically significant differences (P0.05) when compared to baseline control using one-way ANOVA followed Fisher’s protected least squares difference test.
“After continuous daily simvastatin administration, the baseline rCBF was up 25%. L-arginine, but not saline injections, significantly increased rCBF above the simvastatin baseline. In the interval of 10-15 minutes, there was a marked elevation. The maximum elevation was seen at 15-20 minutes and was 29-31% higher than baseline. These increases lasted for 20 minutes more than after L-arginine alone. Statistics did not show statistically an increase in L-arginine’s maximum response when simvastatin was present. The simvastatin-treated mice had a longer response to L-arginine. The simvastatin-treated mice had a greater increase in blood flow during the 30–40 minute time period (P0.05).
“TABLE 1nMABP heart rate, PaO2, PACO2,nGroup n) mmHg pH mmHg and mmHgnwild+ saline ((6)nbaseline 954.7 +/? 3.4 543 +/? 20 7.36 +/? 0.02 154 +/? 13 35.8 +/? 2.0n0-5 min 94.8+/? 3.3 549 +/? 19n10-15 minutes 96.2 +//? 3.2 548 +/? 16n20-25 min 95.8+/? 3.1 550 +/? 16n35-40 min 96.5 2.9 547 +/? 15nafter Infusion 7.35 +/- 0.02 180 +/? 5 33.4 +/? 1.8\nwild + L-arginine (7)\nbaseline 92.9 +/? 3.5* 545 +/? 11 7.40 +/? 0.02 127 +/? 9 39.2 +/? 2.0n0-5 min 92.7+/? 3.5 548 +/? 11n10-15 minutes 94.6 +//? 3.5 561 +/? 9n20-25 min 93.3+/? 3.4 554 +/? 10n35-40 min. 89.9 +//? 3.3 533 +/? 9nafter Infusion 7.32 +/- 0.03 169 +/? 5# 36.4 +/? 1.7neNOS null+ L-narginine ((4)nbaseline 16.3 +/? 9.7 618 +/? 9 7.40 +/? 0.03 15 +/? 4 36.4 +/? 1./n0-5 minutes 115.8+/621 +//? 8./n0-5 min 115.8+/ 621 +/? 7.4 623 +/? 6.4 623 +/- 8.1 630 +/? 13n35-40 min 94.8+/? 8.1 604 +/? 10.2 604 +/- 0.04# 178 +/? 7# 35.8 +/? 2.4\nsim\n(2 mg/kg) + saline\n(3)\nbaseline 88.0 +/? 3.0* 541 +/? 1 7.46 +/? 0.03 159 +/? 19 32.7 +/? 19 32.7 +/? 2.8 543 +/? 3n10-15 minutes 93.3 +//? 3.3 547 +/? 9n20-25 min 95.0+/? 3.5 553 +/? 10n35-40 min 94.0 +//? 4.0 558 +/? 4nafter Infusion 7.41 +/- 0.01 177 +/? 7 32.5 +/? 2.8\nsim (2 mg/kg) + L-\narginine (5)\nbaseline 87.4 +/? 3.1* 505 +/? 9* 7.44 +/? 0.03 144 +/? 15 31.2 +/? 15 31.2 +/? 3.3* 503 +/? 7*n10-15 minutes 92.2 +//? 3.2 507 +/? 6*n20-25 min. 88.4 +/- 3.4* 502 +/? 3.4* 502 +/? 4.2 490 +/? 4*nafter Infusion 7.30 +/- 0.02# 163 +/? 13 34.2 +/? 2.1\nsim\n(20 mg/kg) + L-\narginine (6)\nbaseline 91.7 +/? 2.8* 566 +/? 26 7.44 +/? 0.01 169 +/? 8 32.0 +/? 1.5n0-5 min 91.5+/? 3.7* 571 +/? 26n10-15 minutes 92.7 +/- 4.9 574 +/? 26n20-25 min. 89.7 +/- 4.9* 571 +/? 26n35-40 min 84.8 5.0 551 +/? 21nafter Infusion 7.33 +/- 0.02# 178 +/? 7 32.4 +/? 2.7”
“Example 2”
“A composition containing L-arginine or Simvastatin.”
Mix together “L-Arginine (1. g), Simvastatin (0. g), sucrose (2. g), and purified water(E-Pure 1.5 g). Mix the semisolid mixture until it is homogeneous. It is then dried overnight at 70 C. The semisolid mixture is then ground into particles approximately 1 mm in size. The other half of the particles are then dipped into a 4% solution of benecel in methyl alcohol, and then air-dried.
“These particles are placed into phosphate buffer saline solutions, pH 7.4 at 37 C. The solution is then analyzed at the given time points for L-Arginine.”
“Example 3”
“Another composition containing L-arginine or Simvastatin”
L-Arginine (1 g), Simvastatin (0.25 g), ethylcellulose(Benecel Hercules, 0.3 g), Avicel (FMC), 0.5 g) are all mixed together with purified water (E?Pure). Mix the semisolid mixture until it is homogeneous. It is then dried at 70 C. for four hours. The semisolid mixture is then ground into small particles measuring approximately 1 mm. The other half of the particles are then tumbled in a grinder and gradually coated with a 4% solution in methyl alcohol. The particles are then air dried at 50 C.
Summary for “Increasing cerebral bioavailability for drugs”
The blood-brain barrier prevents compounds from freely circulating in the bloodstream of vertebrates from reaching brain tissue. This barrier protects the brain against external influences. This beneficial role can be detrimental if the drug cannot cross the blood-brain border. It may be impossible to give a drug in sufficient doses to raise the blood levels enough to have a beneficial effect on the brain. This is especially true if the drug being administered to the brain causes side effects that are harmful or undesirable to the rest of the body. This problem can be exacerbated when the condition that requires therapy is associated with reduced blood flow to the brain (e.g., ischemic stroke) or a cardiovascular event that results in decreased blood flow or blood pressure to the brain.
Stroke is the third leading cause of death in developed countries. It can be defined as an abrupt impairment of brain function due to a variety pathologic changes. Ischemic strokes account for approximately 80% of strokes. This is due to restricted blood flow. Patients suffering from strokes may benefit from an increase in blood flow and increased delivery of anti-stroke or neuroprotective drugs.
The pathophysiology and causes of cerebral ischaemia have been linked to both apoptotic and excitotoxic mechanisms. MK-801 is a non-competitive NMDA-channel blocker and glutamate antagonist that protects the brain of rats from ischaemic damage. It is also the prototype for neuroprotective drugs that increase resistance to ischemic injuries. Clinical trials of neuroprotectants failed because inadequate blood (brain level) was not possible or because they were toxic.
“Reduced cerebral blood flow could be a factor in the uptake and penetration of lipophilic drugs such as MK-801 into brain tissue. The uptake of these drugs is severely affected by ischemia and reduced blood flow. Stroke can reduce the possibility of reaching therapeutically appropriate brain levels of neuroprotective drug. It is therefore highly desirable to be able to improve cerebral blood flow and drug delivery, particularly under stroke conditions.
“It is the object of this invention that a method be developed to increase cerebral bioavailability for drugs.”
“It is the object of this invention that a method be developed to increase cerebral bioavailability of drugs as a result of increased cerebral blood flow.”
“Nitric dioxide has been proven to act as a vasodilator in the peripheral vasculature of normal tissue. Surprisingly the inventors discovered that increasing NO levels by endothelial and/or non ecNOS dependent mechanisms, or via the generation of nitric dioxide (eNOS), can affect the vasculature of brain tissue. This causes vasodilation, without any loss in blood pressure. The brain vessels release nitric dioxide, which causes blood flow to increase through the brain tissue. This is independent of blood pressure increases. The present invention uses the blood-pressure-independent increase in blood flow through brain tissue to increase cerebral bioavailability of blood-born compositions.”
The present invention allows for greater cerebral bioavailability and brain NO levels of blood-born compositions. This can be achieved by increasing NO levels by stimulating the production of NO, particularly by administering Larginine. Other agents that increase NO levels independently of ecNOS may also be used. Increased cerebral blood flow means that drugs are also carried into the brain. Increased flow will expose the site of action to more drug molecules. Increased NO production may facilitate administration of drugs not easily absorbed to the brain and/or reduce serum concentrations necessary for desired physiological effects.
“The present invention, in an important embodiment, allows for increased delivery of drugs to brain tissues by administering the drug (also known as the?second agent?) or a?physiologically active compound (or agent). While increasing brain NO levels. This can be achieved by increasing NO levels by stimulating the production of NO, particularly L-arginine. It may also be possible to administer agents that increase NO levels without ecNOS or any combination thereof. These agents should be administered in quantities that increase cerebral blood flow and NO levels. Increased cerebral blood flow means that drugs are also carried into the brain with increased blood flow. Increased flow will expose the site of action to more drug molecules. Increased NO production may facilitate administration of drugs not easily absorbed to the brain and/or reduce serum concentrations necessary for desired physiological effects.
“In one preferred embodiment of this invention, the method for enhancing delivery of a desired composition in brain tissue of an individual comprises introducing the composition substantially concurrently with a blood flow enhancing level of L-arginine.”
“In another preferred embodiment of this invention, this invention provides a means to enhance delivery of a desired component to brain tissue of an person. This involves introducing the composition substantially contemporaneously to a blood flow-enhancing amount L-arginine or a bloodflow-enhancing amount non-ecNOS NO generating system.
Agents such as HMG -CoA reductase and rho -GTPase inhibits and inhibitors of actin cytoskeletal organisation are preferred not to be administered in the methods described in the present invention. They are therefore not included in the compositions of the invention. Preferably, protein kinase C and isoquinolinesulfonyl compound or their derivatives, such as H-7 and H-8, are not administered according to the invention. Cyclosporin-A (Csa-A) may not be administered according to certain embodiments.
“The present invention can be used whenever it is desired to increase cerebral bioavailability for a drug. The term subject is defined as humans, animals other than human primates, and dogs, cats. This invention is therefore useful for therapeutic purposes as well as for research purposes, such as testing in animal models or in vitro models for medical, physiological or metabolic conditions.
“Nitric oxide (NO),” has been identified as an uncommon messenger molecule that plays a variety of physiological roles in the cardiovascular, neurologic, and immune systems. (Griffith T M et. al., J Am Coll Cardiol 1988, 12:797-806). It is responsible for blood vessel relaxation, neurotransmission, and pathogen suppression. NO Synthase produces NO from L-arginine’s guanidino nitrogen. (Moncada, S. and Higgs E., Eur J Clin Invest 1991, 21(4), 361-374). At least three types of NO Synthase are known to exist in mammals. The two calcium-dependent isoenzymes of NO Synthase are expressed in neurons (nNOS), endothelial cells, and macrophages (Type III-ecNOS). Endothelial cell Nitric oxide Synthase is the Type III version of the enzyme that can be found in the endothelium. Its reactivity, different pathways of formation and metabolism can explain the many physiological and pathological consequences of NO.
The cerebral blood flow can be increased to increase the drug’s bioavailability in cerebral areas. The cerebral bioavailability can be increased by increasing cerebral blood flow, according to the present inventors.
The present inventors have shown that brain bioavailability, especially lipophilic drugs can be improved by concurrent administration of L-Arginine and other agents that increase NO production by ecNOS or non-ecNOS. These agents should be administered in quantities that increase cerebral blood flow and NO levels. The compositions of the present invention can also be used prophylactically to reduce the risk of stroke, brain injury, or illness. The present invention also allows for the prophylactic, chronic, and acute administration of compositions. This will increase cerebral uptake of drugs to treat stroke or other brain injuries.
L-arginine is a substrate for endothelial Nitric Ox Synthesise (eNOS). L-arginine administration will increase the mass production of nitric oxygen (NO). L-arginine administration causes cerebral blood flow to increase within minutes. A typical increase in cerebral blood flow can be seen within ten to fifteen minute. The maximum increase may be noticed within twenty to sixty minute. Both the infusion rate and the clearance rate determine the time at which maximum effect occurs. Maximum L-arginine concentration at the end will be achieved if the infusion rate and clearance rate are equal.
The present invention allows for greater cerebral bioavailability by administering the drug at interest and increasing cerebral blood flow through increasing brain NO levels. This may be achieved by increasing NO levels by stimulating the production of NO, particularly L-arginine. Other agents that increase NO levels independently of ecNOS can also be used. These agents should be administered in doses that increase cerebral blood flow and cerebral NO levels. Increased cerebral blood flow means that drugs are also carried into the brain via the blood stream. The site of action will become more vulnerable to drug molecules due to increased flow. The stimulation of increased NO production may facilitate administration of drugs to brains, particularly those that are difficult to introduce to brains. In addition, the serum concentration required to achieve desired physiological effects may be decreased.
There are many ways to measure the NO level in cells or tissues. Endothelial dependent relaxation is one phenotypic measure used in the art. This response is affected by NO levels. A nitric oxygen meter can measure the amount of nitric dioxide in a sample. The techniques described above, along with additional techniques, are all well-known to anyone of average skill in the art.
“The invention allows not only to re-establish normal base-line NO levels, but also allows for an increase in NO levels beyond normal base line levels. Normal base-line levels refer to those found in a normal control group. They are adjusted for age and have no symptoms that would indicate an alteration in nitric oxide levels, such as hypoxic conditions or hyperlipidemia. The assay activity used and the age of the participants will determine the actual level. An abnormal situation, such as stroke, can cause nitric oxide levels to drop below normal levels. In cases of stroke, nitric dioxide levels are depressed below their normal levels. Surprisingly the methods and compositions of the invention can restore normal base-line levels in these abnormal conditions. However, nitric dioxide levels can be raised desirably much higher than normal base-line levels. In the context of this invention, “increasing NO levels” means: This includes both normal NO levels being restored and increasing NO levels beyond normal baseline levels.
The invention includes the treatment of ischemic stroke. Ischemic stroke (ischemic brain infarction) refers to an acute neurologic injury caused by a decrease of blood flow to the brain’s blood vessels. There are two main types of ischemic stroke: thrombotic or embolic.
“An unexpected finding was made regarding the treatment of an ischemic stroke. The invention’s treatment can improve blood flow to the brain even after an ischemic stroke. The present invention led to cerebral blood flow being greater in the treated animals than the control. The increase in nitric dioxide levels is thought to be responsible for the positive results.
“The invention’s most important embodiment is the treatment of subjects at high risk for ischemic stroke. Subjects with an abnormally high risk of having an ischemic stroke as defined herein are those who have been identified by conventional medical practice as having known stroke risk factors or increased risk of developing cerebrovascular events. The risk factors for cardiac disease are similar to those associated with stroke. The primary risk factors include hypertension, hypercholesterolemia, and smoking. Other risk factors include atrial fibrillation and recent myocardial injury.
Subjects at abnormally high risk of an ischemic attack include those who are undergoing diagnostic or surgical procedures that could result in emboli being released, lowering blood pressure, or decreasing blood flow to their brains. These procedures may include brain angiography, carotid endarterectomy and brain angiography. Individuals with abnormally high risk of an ischemic attack include those who have a cardiac condition that could cause decreased blood flow to their brains, such as atrial fibrillation or ventrical tachycardia, dilapidation, and any other conditions that require anticoagulation. Individuals with abnormally high risk of an ischemic attack include those who have conditions such as arteriopathy, brain vasculitis, and congenital blood vessel diseases such as CADASIL syndrome. Migraines are also considered to be at increased risk. “In certain embodiments, the subject does not have a high level of hypercholesterolemic, hypertriglyceridemic, or both.
The treatment of stroke according this invention is for stroke victims who have had a stroke. It can also be used as a prophylactic treatment. Prophylactic treatment for short term is recommended for patients who have undergone diagnostic or surgical procedures that could result in emboli release, lowering blood pressure, or decreases in blood flow to their brains. This will reduce injury from any ischemic events. Patients with cardiac conditions, which may result in decreased blood flow to their brains, or conditions directly affecting the brain’s vasculature, should consider long-term or chronic prophylactic therapy. If the treatment is prophylactic, it will be for those who are at an abnormally high risk of having an ischemic stroke. Acute treatment is possible for subjects who have suffered a stroke. The administration of acute treatment to stroke subjects is preferably at the onset or a significant change in symptoms.
“Another important aspect of the invention is the treatment of subjects suffering from a neurodegenerative disorder. “Neurodegenerative disease” is a broad term. Any pathological condition involving neuronal degradation, such as Huntington’s Disease or Parkinson’s Disease (or ALS), is included in the term “neurodegenerative disease”. Preferably, the neurodegenerative condition is Alzheimer’s Disease. Alzheimer’s Disease, a progressive neurodegenerative disorder, is characterised by the death and loss of function of nerve cells in various areas of the brain. This causes cognitive decline such as memory loss and language loss. Although the cause of nerve cell death remains unknown, the cells can be identified by unusual helical protein filaments (neurofibrillary knots) in nerve cells. Also, degeneration is seen in cortical brain regions, particularly the frontal and temporal. The present invention can increase cerebral bioavailability for suitable drugs. This is especially beneficial to patients suffering from neurodegenerative diseases such as Alzheimer’s.
The present inventors have shown that brain bioavailability, especially lipophilic drugs can be improved by concurrent administration of L-Arginine and other agents that increase NO production by ecNOS or non-ecNOS. These agents should be administered in sufficient quantities to increase cerebral blood flow and brain NO levels. The compositions of the invention can also be used prophylactically to reduce the risk of stroke, other brain injuries, or illnesses. The present invention also promotes cerebral uptake of drugs to treat stroke and other brain injuries.
The present invention allows for greater cerebral bioavailability by administering the drug and increasing cerebral blood flow through the increase in NO levels. This may be achieved by increasing NO levels by increasing eNOS production, particularly by administering Larginine. Other agents that increase NO levels independently of ecNOS can also be used. These agents should be administered in sufficient quantities to increase blood flow to the brain and/or NO levels. Increased cerebral blood flow means that drugs are also carried into the brain with increased blood flow. The site of action will become more vulnerable to drug molecules due to increased flow. The stimulation of increased NO production may facilitate administration of drugs to brains, particularly those that are difficult to introduce to brains. In addition, the serum concentration required to achieve desired physiological effects may be decreased.
An agent (preferably Larginine) that increases the ecNOS levels by preexisting and/or non-ecNOS NO generation is administered to the subject who requires enhanced drug delivery. An agent that increases ecNOS by preexisting (preferably L-arginine), and/or a system that does not generate NO may be called a?NO-increasing combination. Each of these agents can be called an ‘NO-increasing agency. These agents could be combined to be called?NO-increasing agent.
“As used in the present application, the terms “NO-increasing cocktail?” a?NO-increasing agent? Agents such as HMG?CoA reductase inhibits, rho?GTPase inducers and inhibitors for actin cytoskeletal order are not included. These agents are described in detail in WO 99/18952, WOM 99/47153 and WOM 00/03746, which have been incorporated herein by reference. The terms “NO-increasing cocktail” and “NO-increasing agent(s)” are used throughout this application. a?NO-increasing agent? Do not contain protein kinase C inhibitors, Cyclosporin-A (Csa-A), or Isoquinolinesulfonyl compound or their derivatives, such as H-7 or H-8.
The NO-increasing agent/mixture is administered substantially simultaneously with the second agent (the drug whose delivery will be improved). The preferred embodiment of the NO-increasing cocktails includes L-arginine, at least one nonecNOS NO generating system, and L-arginine.
“The components of the NO -increasing cocktail should be administered substantially simultaneously with each other. As we will see, “substantially contemporaneous administration” is a term that can be used to describe a combination of components. It should be understood broadly and includes many types of administration. Referring to the components of NO-increasing cocktails, “substantially contemporaneously?” This means that the components are administered in such a way that they work together to produce a significant increase in NO levels in the subject or in particular tissues.
To be considered a “NO-increasing cocktails,” agents not only need to be administered in one form or one unitary dose, but also in multiple doses. “NO-increasing cocktails” does not mean that agents used in the present invention must be administered in a single formulation or in one unitary dosage.
“To stimulate higher NO production by ecNOS an?ecNOS activating ingredient (or agent) is required. is administered to the subject who requires enhanced drug delivery. Agents that increase NO production through ecNOS Agents that increase NO production through preexisting ecNOS. These terms can also be used interchangeably to denote ecNOS activating agent. These agents are also known as agents which increase NO production via preexisting EcNOS. This does not mean that these agents can’t also increase NO production of ecNOS produced during or after administration. Instead, the term “preexisting” is used. This is to show that these agents don’t increase or downregulate the expression ecNOS.
The activating component must be administered concurrently with the drug whose delivery is being enhanced. An eNOS substrate such as L-arginine is a preferred component that activates eNOS. This increases NO production. Alternative eNOS activating components include cofactors of eNOS, such as NADPH or tetrahydrobiopterin.”
“Compounds that increase NO production by preexisting ecNOS can do so through a variety of mechanisms. L-arginine and other ecNOS substrates increase NO production by mass action. NADPH and Tetrahydrobiopterin are cofactors that increase NO production by increasing the ability for ecNOS catalyze the substrate to NO conversion. These ecNOS substrates, e.g. L-arginine and cofactors (e.g. NADPH, Tetrahydrobiopterin etc.) may be used as ecNOS substrates. Natural or synthetic, they can be either natural or artificial. “Compounds that increase NO production by preexisting NOS can act in a cooperative, additive, or synergistic manner with agents that increase NO levels through non-ecNOS dependent mechanisms (i.e. non-ecNOS no-generating systems).
L-arginine is a substrate for endothelial Nitric Ox Synthesise (ecNOS). L-arginine administration will increase the mass production of nitric oxygen (NO). L-arginine administration causes cerebral blood flow to increase within minutes. A typical increase in cerebral blood flow can be seen within ten to fifteen minute. The maximum increase can occur within twenty to sixty minute. Both the infusion rate and the clearance rate determine the time at which maximum effect occurs. Maximum L-arginine concentration at the end will be achieved if the infusion rate and clearance rate are equal.
“EcNOS activating components” is the term used in the present application. HMG-CoA reductase inhibits, rho?GTPase inducers and inhibitors of actin-cytoskeletal organization are not included. These agents are described in detail in WO 99/18952, WOM 99/47153 and WOM 00/03746, which have been incorporated herein by reference. The term “ecNOS activating constituent” is used throughout this application. Does not include protein kinase C inhibits, cyclosporin A (Cs-A), isoquinolinesulfonyl compound or their derivatives. These are not administered according to the invention and are not included within the compositions according the invention.
When administered to biological systems, “NO donors” are compounds that release or produce NO, or have NO-related activity. Feelisch (1998) Naunyn Schmiedebergs Arch Pharmacol 358(1): 113-22. There are many examples of NO donors that are well-known and will be discussed in detail below. Subjects can also increase NO levels by inhaling NO.
“Non-ecNOS NO generating devices directly increase NO levels in a subject, tissue, or cell without relying upon ecNOS and other Nitric Oxide synthases. The invention defines “non-ecNOS” NO-generating systems as: Compounds that, when administered to a subject increase NO levels without relying upon ecNOS. Inhalation of NO can be used to administer NO directly to the subject. A patient may also receive NO via administration of NO donors. “Non-ecNOS NO generating systems” includes both NO and NO donors.
“The term “non-ecNOS no-generating systems” is used throughout this application. HMG-CoA reductase inhibits, rho?GTPase inhibitions and inhibitors for actin cytoskeletal organisation are not included. These agents are described in detail in WO 99/18952, WOM 99/47153 and WOM 00/03746, which have been incorporated herein by reference. The term “non-ecNOS no-generating systems” is used throughout this application. Does not include protein kinase C inhibitors (Cs-A), cyclosporin A (Cs-A), isoquinolinesulfonyl compound or their derivatives. These are not administered according to the invention and are not included within the compositions according the invention.
“It is possible to optimize routinely the administration of inhaled NO for the use in the compositions and methods of the present invention.” The properties and activities of NO inhaled are well-known. Administration of inhaled NO to human subjects is described at least in Hoeper, et al., Abman, et al., Carrier, et al., Kinsella, et al., and Kuhlen, et al. See, e.g., Hoeper, et al. (1999) Respir Med. 93(1): 62-4; Abman, et al. (1994) J. Pediatr. 124(6): 881-8; Carrier, et al. (1999) 18(7): 664-7; Kinsella, et al. (1993) J. Pediatr. 122(5Pt 1): 803-6; Kuhlen, et.al. (1999) Intensive Care Med. 25(7): 752-4. The texts of these publications are incorporated by reference herein.
Multiple groups have described the chemical kinetics and methods of reducing NO2 buildup in systems for inhalation of NO2. See, e.g., Tsukahara, et al. (1999) Nitric Oxide (3): 191-8. Lindberg, et. al. (1998) Br. J Anaesth 80(2) 213-7. The texts of these publications are included herein as a reference. Multiple groups have also provided detailed descriptions of techniques for monitoring and delivery of inhaled NO. See, e.g., Kirmse, et al. (1998) Chest 113(6): 1650-7; Shibata (1996) Acta Paediatr Jpn 38(2): 143-6; Young, et al. 1996 Intensive Care Med. 22(1):77-86; Hess, and al. Respir Care Clin N Am 3, 3(3), 371-410. The texts of these publications are included herein as a reference. The art describes technical considerations such as NO gas concentrations. See, e.g., Kinsella, et al. (1999) Curr Opin Pediatr 11(2): 121-5; Foubert, et al., (1999) Anaesthesia 54(3): 220-5; Breuer, et al. (1997) Eur. J. Pediatr. 156(6): 460-2; Moon, et al. (1997) Biomed Instrum Technol 31.2: 164-8; Hart (1999) Chest 115.5: 1407-17. The texts of these publications are included herein as a reference.
“As used herein, the term ‘NO donors? A large group of molecules that have varying properties but all release or produce NO, or are related to NO, when applied to biological systems. Feelisch (1998) Naunyn Schmiedebergs Arch Pharnacol 358,(I): 113-22. The text of that publication is included herein as a reference. These compounds are well-known in art. Examples of NO donors include nitroglycerin, nitric oxide/nucleophile adducts (NONOates), including diethylamine/NO complex sodium (Dea/NO) and spermine/NO complex sodium; S-nitrosothiols, also called NO+ equivalents, such as S-nitroso-L-glutathione (GSNO) and S-nitroso-N-acetyl-D,L-penicillamine (SNAP); nitrosylated proteins, such as nitrosylated bovine serum albumin (BSA). See, e.g., Ewing, et al., (1997) J. Pharmacol. Exp. Ther. 283(2):947-54; Vidwans, et al. (1999). J. Neurochem 72(5) : 947-54; Vidwans, et al. SPM-5185, an organic cysteine-containing drug, undergoes a biotransformation process that releases NO when it is exposed to physiological conditions. See, e.g., Vinten-Johansen, et al. (1995) Int. J Cardiol. 50(3): 273?81. The text of that publication is incorporated by reference herein. Another NO donor that has been therapeutically used in humans is sodium nitrosprusside. See, e.g., Thomas, et al. (1999) Neurosurgery 44(1):48-57, 57-8; Thomas, et al. (1999) Stroke (30(7): 1409-16. The texts of these publications are included herein by reference. The heterocyclic NO-releasing compound, which includes mesionic heterocycles such as SIN-1 and heterocyclic NOs such as furoxane carbboxamides, are other examples of NO donors. See, e.g., Schonafinger (1999) Farmaco 54(5): 316-20 and Hou, et al., (1999) Curr. Pharm. Des. 5(6): 417-441. The texts of these publications are incorporated by reference. U.S. Patent. Describes additional NO donors. No. 5,910,316 to Keefer, et al., U.S. Pat. No. 5,525,357 to Keefer, et al., U.S. Pat. No. Loscalzo, et al. and U.S. Pat. No. No.
It is routine optimization for a skilled worker in the art to choose NO donors that are suitable for the compositions and methods according to the invention. A skilled worker in the arts can also routinely optimize dosages and administration methods to suit the compositions and techniques of the invention. The activities and properties of NO donors have been well-known. Schmidt et al. have created a mathematical model to predict the NO concentrations of donor compounds over time. Schmidt, et al. Schmidt, et al. Morley and Keefer discuss in depth NONOates. Kal, et. al. describes in detail how nitroglycerin is administered to patients. Morley, et al., (1993) J. Cardiovasc. Pharmacol. 22 Suppl. 7: S3-9; Kal, et al. (1999) Anesth. Analg. Analg.
“Estrogens, ACE inhibitors and other hormones can also increase NO levels. While estrogens and ACE inhibitors can be used in the compositions and methods according to the invention, they are not included under the term?agents that increase NO production by preexisting ecNOS? ?NO-increasing compound,? ?NO-increasing cocktail,? ?non-ecNOS NO-generating system,? or ?ecNOS activating component.? Estrogens can be described as a specific group of molecules that are well-known to those with ordinary skill in the field. We won’t go into detail about them. They share high levels of structural similarity. ACE inhibitors have also been well characterized. However, they don’t always share structural homology.
Angiotensin converting enzyme (or ACE) is an enzyme that catalyzes angiotensin I and II conversions. ACE inhibitors are amino acids, peptides (including di and tri peptides), and antibodies to ACE. They interfere in the renin?angiotensin systems by inhibiting the activity ACE and thereby reducing/eliminating the formation of the pressor substance angiotensin 2. ACE inhibitors are used medically to treat hypertension and congestive heart disease, as well as myocardial injury, myocardial damage, renal disease, and congestive heart failure. ACE inhibitors are acylmercapto, mercaptoalkanoyl proslines like captopril (U.S. Patent. No. No. No. No. No. 4,374,829), lisinopril (U.S. Pat. No. 4,374,829), quinapril (U.S. Pat. No. No. No. No. No. No. No. No. No. 4,410,520), phosphinylalkanoyl proslines such as fosinopril. (U.S. Pat. No. No.
“In some embodiments, the second drug (i.e. the drug that is sought to enhance delivery) is administered to a subject suffering from a condition. The second agent is administered in sufficient amounts to treat the condition. This increases blood flow to the subject’s tissues.
The second agent can be any pharmacological or diagnostic compound. Agents with a brain-specific site of action are preferred second agents. These agents can be analgesics, anesthetics, anti-andrenergic agents, amino acids and antagonists, as well as antidote (anti-anxiety agent), anticolvunsant/antidepressant/antihypertensive/antihypertensive/antifibrinolytics, antihypertensive/antihypertensive/antiparkinsonian agent, antiobessional agent or antiparkins, neuromuscular block, neuroprotective, and neuromuscular blockers, neuromuscular blockers, and neuroprotective, serotonin agonists, and calcium-channels. NMDA antagonist, post-stroke and post-head trauma treatment, psychotropic, sedative, sedative-hypnotic, selective serotonin uptake inhibitor, serotonin inhibitor, tranquilizer, and treatment of cerebral ischemia, calcium channel blockers, free radical scavengers-antioxidants, GABA agonists, glutamate antagonists, AMPA antagonists, kainate antagonists, competitive and non-competitive NMDA antagonists, growth factors, opioid antagonists, phosphatidylcholine precursors, serotonin agonists, sodium- and calcium-channel blockers, and potassium channel openers.”
“In addition to the foregoing brain-specific categories of agents, examples of categories of other pharmaceutical agents that can be used as second agents include: adrenergic agent; adrenocortical steroid; adrenocortical suppressant; alcohol deterrent; aldosterone antagonist; amino acid; ammonia detoxicant; anabolic; analeptic; analgesic; androgen; anesthesia, adjunct to; anesthetic; anorectic; antagonist; anterior pituitary suppressant; anthelmintic; anti-acne agent; anti-adrenergic; anti-allergic; anti-amebic; anti-androgen; anti-anemic; anti-anginal; anti-anxiety; anti-arthritic; anti-asthmatic; anti-atherosclerotic; antibacterial; anticholelithic; anticholelithogenic; anticholinergic; anticoagulant; anticoccidal; anticonvulsant; antidepressant; antidiabetic; antidiarrheal; antidiuretic; antidote; anti-emetic; anti-epileptic; anti-estrogen; antifibrinolytic; antifungal; antiglaucoma agent; antihemophilic; antihemorrhagic; antihistamine; antihyperlipidemia; antihyperlipoproteinemic; antihypertensive; anti-infective; anti-infective, topical; anti-inflammatory; antikeratinizing agent; antimalarial; antimicrobial; antimigraine; antimitotic; antimycotic, antinauseant, antineoplastic, antineutropenic, antiobessional agent; antiparasitic; antiparkinsonian; antiperistaltic, antipneumocystic; antiproliferative; antiprostatic hypertrophy; antiprotozoal; antipruritic; antipsychotic; antirheumatic; antischistosomal; antiseborrheic; antisecretory; antispasmodic; antithrombotic; antitussive; anti-ulcerative; anti-urolithic; antiviral; appetite suppressant; benign prostatic hyperplasia therapy agent; blood glucose regulator; bone resorption inhibitor; bronchodilator; carbonic anhydrase inhibitor; cardiac depressant; cardioprotectant; cardiotonic; cardiovascular agent; choleretic; cholinergic; cholinergic agonist; cholinesterase deactivator; coccidiostat; cognition adjuvant; cognition enhancer; depressant; diagnostic aid; diuretic; dopaminergic agent; ectoparasiticide; emetic; enzyme inhibitor; estrogen; fibrinolytic; fluorescent agent; free oxygen radical scavenger; gastrointestinal motility effector; glucocorticoid; gonad-stimulating principle; hair growth stimulant; hemostatic; histamine H2 receptor antagonists; hormone; hypocholesterolemic; hypoglycemic; hypolipidemic; hypotensive; imaging agent; immunizing agent; immunomodulator; immunoregulator; immunostimulant; immunosuppressant; impotence therapy adjunct; inhibitor; keratolytic; LNRH agonist; liver disorder treatment; luteolysin; memory adjuvant; mental performance enhancer; mood regulator; mucolytic; mucosal protective agent; mydriatic; nasal decongestant; neuromuscular blocking agent; neuroprotective; NMDA antagonist; non-hormonal sterol derivative; oxytocic; plasminogen activator; platelet activating factor antagonist; platelet aggregation inhibitor; post-stroke and post-head trauma treatment; potentiator; progestin; prostaglandin; prostate growth inhibitor; prothyrotropin; psychotropic; pulmonary surface; radioactive agent; regulator; relaxant; repartitioning agent; scabicide; sclerosing agent; sedative; sedative-hypnotic; selective adenosine Al antagonist; serotonin antagonist; serotonin inhibitor; serotonin receptor antagonist; steroid; stimulant; suppressant; symptomatic multiple sclerosis; synergist; thyroid hormone; thyroid inhibitor; thyromimetic; tranquilizer; treatment of amyotrophic lateral sclerosis; treatment of cerebral ischemia; treatment of Paget’s disease; treatment of unstable angina; uricosuric; vasoconstrictor; vasodilator; vulnerary; wound healing agent; xanthine oxidase inhibitor.”
“Throughout this application by terms such as “substantially contemporaneous Administration”,? ?co-administration,? ?substantially contemporaneously,? ?substantially concurrently? It is intended that all of the administered compounds are administered to the subject in a relative order so that they may exert an additive, or even synergistic effect on the subject. On increasing NO levels or delivering a second agent via increased blood flow to a tissue. These terms can be interchanged.
“Substantially contemporaneous Administration? “?Substantially contemporaneous administration” refers to administration of a combination of drugs for increasing NO levels (as discussed herein), such that cerebral blood flow is increased while the second medication is in sufficient serum concentration (i.e. enough to allow the second drug have a physiological effect).
“With respect to NO-increasing agent and components of the NO?increasing cocktail??substantially contemporaneously?” This means that the timing of administration of components is coordinated in such a way that a significant increase in NO levels is achieved in the subject or in particular tissues.
“Substantially simultaneous administration?” This includes administration of agents (both NO -increasing agent and second agents) in one formulation or as a unitary dose. Substantially simultaneous administration This includes agents administered in different dosage formats, formulations, and at different times. As long as the criteria are met for substantially simultaneous administration (as noted above),
“For example, a combination of drugs for increasing NO levels and the second drug could be prepared for intravenous administration. Infusion in one pharmaceutical composition. This allows for simultaneous infusion of a combination that increases NO levels and the second drug. It is preferable that the drug to which the delivery is being increased is in the bloodstream while NO levels are increasing.
The two components can be combined in one oral composition if the second drug is absorbed into bloodstream at a rate similar to that of an oral combination for increasing NO levels (as discussed herein). The pharmacokinetics for the second drug may not be as effective for several hours. If this happens, a combination for increasing NO levels (as discussed herein) can be administered in substantially contemporaneous fashion. This means that the blood flow will increase once the serum concentrations of both drugs have been reached. A combination of drugs for increasing NO levels, as described herein, that are substantially contemporaneous in administration to another drug, whether one or both of them are administered via a nasal, topal, or rectal route or intramuscularly or undercutaneously, is routine for those who are skilled in pharmacology or clinical medicine.
Effective amounts are given to agents (preferably Larginine), which increase NO production by preexisting NO generating systems and/or ecNOS NO generating system. An effective amount is generally any amount that causes an increase in blood flow to or from the brain. This is usually a amount that is effective in increasing NO levels in the brain.
The second or additional agents can also be administered in effective doses. An effective amount of any pharmaceutical preparation is the amount that produces the desired response, whether it’s taken alone or in combination with other agents. It may be a temporary slowing down of the disease’s progression, but it is more effective if the disease is stopped permanently. Routine methods can monitor such results.
The effectiveness of a second agent will depend on how it is administered. The effective amounts of second agent are known or can be easily determined by those skilled in clinical medicine and pharmacology. The present invention aims to decrease the amount of second agents required to achieve a desired effect. Therefore, the effective amounts can be modified by administering the second agent according to the methods of this invention. The new effective amounts can be determined by routine experimentation of those who are skilled in clinical medicine and pharmacology.
The effective doses of both the NO-increasing and second agents will vary depending on the patient’s condition and their individual parameters, including their age, weight and size, as well as the type of concurrent therapy and other factors that the health practitioner has access to. Some forms of administration such as intravenous injections can result in lower doses. If a subject’s response is not sufficient at the initial doses, they may need to be administered higher doses or a more localized route. To achieve the appropriate systemic levels, multiple doses are possible per day.
“It is preferred that the maximum dose of NO-increasing agent or cocktail be used. This is the safest dose according to sound medical judgement. However, those with ordinary skill in the arts will understand that patients may request a lower or more tolerable dose for psychological or medical reasons.
“The agents that increase NO production by preexisting ecNOS, NO, NO donors and other compounds useful according to the invention may be combined, optionally, with a pharmaceutically-acceptable carrier. The term ?pharmaceutically-acceptable carrier? The term “pharmaceutically-acceptable carrier” is used herein to refer to any combination of solid or liquid fillers, diluting agents or encapsulating substances that are suitable for administration into a person. “Carrier” is a term that refers to an organic or inorganic ingredient. The term “carrier” refers to an organic or inorganic component, natural or synthetic, that is used to combine the active ingredient for the purpose of application. All components of the pharmaceutical compositions are also capable of being mixed with the molecules of this invention and with each other in a way that does not substantially impair their pharmaceutical efficacy.
“The pharmaceutical compositions could contain appropriate buffering agents such as acetic acid, citric acid and salt; boric acids in salt; or phosphoric acid.
“The pharmaceutical compositions may also contain, optionally: benzalkonium chloride, chlorobutanol, parabens, and thimerosal.”
There are many routes of administration. There are many options available. The mode of administration will depend on the drug being used, the severity of the condition, and the dose required to achieve therapeutic efficacy. Medically acceptable means that the methods can be used to produce effective levels of active compounds with no clinically unacceptable side effects. These modes of administration include oral, rectal and topical, nasal, interdermal or parenteral. Parenteral is a broad term. It can be subcutaneous, intravenous or intramuscular. For prophylaxis and long-term treatment, intravenous and intramuscular routes may not be suitable.
The pharmaceutical compositions can be conveniently presented in a unit dosage form. They may also be prepared using any of the well-known methods in pharmacy. All methods involve the addition of the active agent(s), or any combination thereof, to a carrier. The compositions are generally prepared by bringing the active compounds into contact with a liquid carrier or a finely divided solid carrier. If necessary, the product can be shaped.
“Compositions that are suitable for oral administration can be presented in discrete units such as tablets, capsules, or lozenges. Each unit contains a predetermined amount. You can also make suspensions from aqueous or non-aqueous liquids, such as syrups, elixirs or emulsions.
“Compositions that are suitable for parenteral administration consist of a sterile, aqueous preparation containing a NO-increasing or cocktail. It should be preferably isotonic with blood of the recipient. This composition may also contain the second or additional agents. They may also be administered separately, but in a substantially parallel fashion. The aqueous preparation can be prepared according to well-known methods, using appropriate dispersing or wetting agent and suspending agents. The sterile injectable preparation also may be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example, as a solution in 1,3-butane diol. Water, Ringer’s solution and isotonic chloride solution are acceptable solvents. Sterile, fixed oils can also be used as a solvent and suspending medium. Any bland, unscented oil can be used for this purpose, including synthetic mono- or di-glycerides. You can also use fatty acids like oleic acid in the preparation injectables. Carrier formulation suitable for oral, subcutaneous, intravenous, intramuscular, etc. Remington’s Pharmaceutical Sciences, Mack Publishing Co. Easton, Pa. has the administrations.
“Other delivery methods include delayed release, time-release and sustained release. These systems allow for the patient to be administered once and then stopped. This increases convenience for both the subject as well as the physician. There are many types of release delivery systems that are known to anyone with ordinary skill in the arts. They include polymer base systems such as poly(lactide-glycolide), copolyoxalates, polycaprolactones, polyacrylates, polyesteramides, polyorthoesters, polyhydroxybutyric acid, and polyanhydrides. U.S. Pat. describes microcapsules containing the above-mentioned polymers that contain drugs. No. 5,075,109. Non-polymer delivery systems include: lipids, including cholesterol esters and neutral fats like mono-di and tri-glycerides, hydrogel release systems, cellusics, sylastic system; peptide-based systems; wax coatings and compressed tablets with conventional binders or excipients. Examples include, but not limited to: (a. erosional systems where the active compound is contained within a matrix like those described in U.S. Pat. Nos. Nos. Nos. Nos. Pump-based hardware delivery systems are also available, some of which can be adapted for implant.
A long-term sustained-release implant might be desirable. The term “long-term release” is used herein to mean that the implant has been constructed and designed to deliver therapeutic levels of active ingredient for at most 30 days and preferably 60. Long-term sustained-release implants are well-known to anyone with ordinary skill in medicine and include some of these release systems.
Pre-packaged combinations of NO increasing agent or agents and NO-increasing cocktail, second agents, or any combination thereof, are preferred embodiments. Combinations of NO-increasing agent/ agents, NO-increasing cocktail, second agents and any combination thereof that are prepackaged using “Blow/Fill/Seal” are even more preferred. technology, as described below.”
“Blow-Fill Seal technology” is a manufacturing process that allows for simultaneous filling and sealing of containers using one machine. Sometimes called form-fill-seal. This process has many advantages over traditional aseptic filling preformed plastic containers (or other types) by eliminating the need for multiple steps. It can also be performed in one machine in a sterile environment and requires very little operator intervention. (J. R. Sharp.?Manufacture Sterile Pharmaceutical Products Using ‘Blow-Fill? Seal? Technology?, Pharm. J., 239, 106 (1987). F. Leo?Blow/Fill/Seal Aseptic Package Technology in Aseptic Pharmaceutical Tech for the 1990’s?. Interpharm Press. Prairie View, Ill. 1989, pp. 195-218).”
The manufacturing process is broken down into several stages. In stage 1, the polyethelene resin undergoes high pressure and temperature, and is then extruded continuously to form a tube. This is known as a parison. The mold is closed when the tube is the right length and the parison cut. The top of the parison can be held in place by securing the bottom. The mold is then moved to the position where it can be filled with sterilized air. The second stage involves the inserting the blow-fillnozzle into the parison, until it seals with the mold’s neck. By blowing compressed air through the parison and expanding it against the walls of a mold cavity, the container is created. The compressed air is then ejected from the container, and a sterile product is poured into the container via the fill nozzle. The nozzle can be retracted back to its original position after the container has been filled. Semi-molten is reached at this stage of the cycle (stage 3). To form the top, separate sealing molds are used to seal the container. The mold is then opened in stage 4. The mold is removed from the machine and the container is filled, sealed, and returned to the origin point to begin the next cycle. You can add additional products to the container anytime during or prior to use through an injection port.
“A preferred embodiment contains a NO-increasing cocktail or agent and a second agent. It is packaged in a Blow/Fill/Seal containers according to the methods herein. Another preferred embodiment is that a NO-increasing cocktail or agent is prepackaged into a Blow/Fill/Seal containers according to the methods herein. A second agent or agents can be injected into this container before or during administration to patients. The second agent may be administered by another method such as injection, oral administration (sublingual administration), inhalation or the like.
“In all of the above embodiments it is preferable that L-arginine be the NO-increasing agents or that L-arginine is the NO-increasing cocktail.”
“In such prepackaging embodiments the NO-increasing agents or cocktails included in these prepackaging embodiments will preferably be sufficient to increase blood flow and NO levels in the brain of the subject. Prepackaged formulations will contain sufficient second agents to achieve the desired effect. It is especially important that the amount of the second agent be adjusted to account for the increased blood flow caused by the NO-increasing agents or cocktails. The effectiveness of each second agent will differ.
“It will be obvious to the skilled artisan, that the considerations in formulating pharmaceutical preparations, and determining the modes of administration for non-ecNOS-generating systems, are similar to those involved in such formulation, determination for agents which upregulate the expression of ecNOS, and compounds which increase NO production by preexisting, ecNOS.”
“Publications WO 99/18952, WO99/47153, WO 00/03746, and WO 00/03746, provide more detailed information about endothelial nitricoxid synthase and their regulation, as well methods of formulating compounds that affect eNOS such as L-arginine and the physiologically active compositions. These publications are herein incorporated as reference.”
“EXAMPLES”
Below are a few examples that will help you better understand the invention. These Examples are only examples and do not limit the scope of the invention.
“Example 1”
“Effects of L-arginine upon Cerebral Blood Flow.”
Infusion of L-arginine at 300 mg/kg i.v. caused moderate (10%) and variable elevations (RCBF), after infusion in several preliminary studies (n=4, data unpublished). The present experiments showed that simvastatin (2 mg/kg) or 450 mg/kg of saline were infused at a steady rate of 100 microliter/kg/min for 15 minutes to wild type mice, mutant mice lacking endothelial oxide synthase, and mice who had been given simvastatin (2 mg/kg) daily. Regional cerebral blood flow (rCBF) was monitored by laser-Doppler flowimetry in groups of urethane-anesthetized, ventilated mice. Other physiological variables, such as mean arterial blood pressure (MABP), heart beat, blood pH, and PaO2 were also observed in the mice.
“Results”
“Physiological variables during laser-Doppler flowimetry in urethane-anesthetized ventilated wild type, simvastatin-treated and eNOS null mice infused with L-arginine or saline are shown in Table 1. In parenthesis, the number of mice in each group are shown. The values are reported as the mean +/-? SEM. SEM. MABP is the mean arterial blood pressure. Sim denotes mice who have been chronically administered simvastatin.
“There were no differences between the groups in heart rate and mean arterial blood pressure during observation, but these values were higher in eNOS-null mice, as previously reported. PaCO2 values did not differ between the two time points of all groups or between-groups, but pH values were lower after L-arginine infusion.
“rCBF Response for L-arginine”
“FIG. “FIG. In parenthesis, the number of mice in each group are indicated. An error bar denotes standard error of mean (SEM) and an asterisk(*) denotes statistically significantly difference (P0.05) when compared to baseline control using one-way ANOVA followed Fisher’s protected least squares difference test.
“L-arginine infusion (450 mg/kg, i.v.) As shown in FIG. 1 (FIG. 1). At 5-10 minutes, the RCBF increased and reached statistical significance 10-15 minutes after infusion. Maximum values reached at 20-25 minutes were 26% higher than the control level, and then they decreased to their normal levels. L-arginine, however, did not increase the rCBF of eNOS null mice. These mutants had values ranging from?4 up to +5% over the 40-minute recording period. In wild-type mice, saline infusion did not significantly increase rCBF.
“rCBF Response To L-arginine Plus Simvastatin.”
“FIG. “FIG. In parenthesis, the number of mice in each treatment group is indicated. sim denotes simvastatin. An error bar denotes SEM, and an asterisk (“*”) denotes statistically significant differences (P0.05) when compared to baseline control using one-way ANOVA followed Fisher’s protected least squares difference test.
“After continuous daily simvastatin administration, the baseline rCBF was up 25%. L-arginine, but not saline injections, significantly increased rCBF above the simvastatin baseline. In the interval of 10-15 minutes, there was a marked elevation. The maximum elevation was seen at 15-20 minutes and was 29-31% higher than baseline. These increases lasted for 20 minutes more than after L-arginine alone. Statistics did not show statistically an increase in L-arginine’s maximum response when simvastatin was present. The simvastatin-treated mice had a longer response to L-arginine. The simvastatin-treated mice had a greater increase in blood flow during the 30–40 minute time period (P0.05).
“TABLE 1nMABP heart rate, PaO2, PACO2,nGroup n) mmHg pH mmHg and mmHgnwild+ saline ((6)nbaseline 954.7 +/? 3.4 543 +/? 20 7.36 +/? 0.02 154 +/? 13 35.8 +/? 2.0n0-5 min 94.8+/? 3.3 549 +/? 19n10-15 minutes 96.2 +//? 3.2 548 +/? 16n20-25 min 95.8+/? 3.1 550 +/? 16n35-40 min 96.5 2.9 547 +/? 15nafter Infusion 7.35 +/- 0.02 180 +/? 5 33.4 +/? 1.8\nwild + L-arginine (7)\nbaseline 92.9 +/? 3.5* 545 +/? 11 7.40 +/? 0.02 127 +/? 9 39.2 +/? 2.0n0-5 min 92.7+/? 3.5 548 +/? 11n10-15 minutes 94.6 +//? 3.5 561 +/? 9n20-25 min 93.3+/? 3.4 554 +/? 10n35-40 min. 89.9 +//? 3.3 533 +/? 9nafter Infusion 7.32 +/- 0.03 169 +/? 5# 36.4 +/? 1.7neNOS null+ L-narginine ((4)nbaseline 16.3 +/? 9.7 618 +/? 9 7.40 +/? 0.03 15 +/? 4 36.4 +/? 1./n0-5 minutes 115.8+/621 +//? 8./n0-5 min 115.8+/ 621 +/? 7.4 623 +/? 6.4 623 +/- 8.1 630 +/? 13n35-40 min 94.8+/? 8.1 604 +/? 10.2 604 +/- 0.04# 178 +/? 7# 35.8 +/? 2.4\nsim\n(2 mg/kg) + saline\n(3)\nbaseline 88.0 +/? 3.0* 541 +/? 1 7.46 +/? 0.03 159 +/? 19 32.7 +/? 19 32.7 +/? 2.8 543 +/? 3n10-15 minutes 93.3 +//? 3.3 547 +/? 9n20-25 min 95.0+/? 3.5 553 +/? 10n35-40 min 94.0 +//? 4.0 558 +/? 4nafter Infusion 7.41 +/- 0.01 177 +/? 7 32.5 +/? 2.8\nsim (2 mg/kg) + L-\narginine (5)\nbaseline 87.4 +/? 3.1* 505 +/? 9* 7.44 +/? 0.03 144 +/? 15 31.2 +/? 15 31.2 +/? 3.3* 503 +/? 7*n10-15 minutes 92.2 +//? 3.2 507 +/? 6*n20-25 min. 88.4 +/- 3.4* 502 +/? 3.4* 502 +/? 4.2 490 +/? 4*nafter Infusion 7.30 +/- 0.02# 163 +/? 13 34.2 +/? 2.1\nsim\n(20 mg/kg) + L-\narginine (6)\nbaseline 91.7 +/? 2.8* 566 +/? 26 7.44 +/? 0.01 169 +/? 8 32.0 +/? 1.5n0-5 min 91.5+/? 3.7* 571 +/? 26n10-15 minutes 92.7 +/- 4.9 574 +/? 26n20-25 min. 89.7 +/- 4.9* 571 +/? 26n35-40 min 84.8 5.0 551 +/? 21nafter Infusion 7.33 +/- 0.02# 178 +/? 7 32.4 +/? 2.7”
“Example 2”
“A composition containing L-arginine or Simvastatin.”
Mix together “L-Arginine (1. g), Simvastatin (0. g), sucrose (2. g), and purified water(E-Pure 1.5 g). Mix the semisolid mixture until it is homogeneous. It is then dried overnight at 70 C. The semisolid mixture is then ground into particles approximately 1 mm in size. The other half of the particles are then dipped into a 4% solution of benecel in methyl alcohol, and then air-dried.
“These particles are placed into phosphate buffer saline solutions, pH 7.4 at 37 C. The solution is then analyzed at the given time points for L-Arginine.”
“Example 3”
“Another composition containing L-arginine or Simvastatin”
L-Arginine (1 g), Simvastatin (0.25 g), ethylcellulose(Benecel Hercules, 0.3 g), Avicel (FMC), 0.5 g) are all mixed together with purified water (E?Pure). Mix the semisolid mixture until it is homogeneous. It is then dried at 70 C. for four hours. The semisolid mixture is then ground into small particles measuring approximately 1 mm. The other half of the particles are then tumbled in a grinder and gradually coated with a 4% solution in methyl alcohol. The particles are then air dried at 50 C.
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